A major hurdle in the development of effective vaccines against pathogens that reside within macrophages, including Mycobacterium tuberculosis, is how to deliver exogenous antigens in a manner that stimulates a protective cellular immune response. Recent investigations involving antisense mutants of M. tuberculosis H37Rv that have diminished production of superoxide dismutase (SOD) and exhibit promising activity as a vaccine prototype suggest that the mechanism of vaccine efficacy may be apoptosis-associated cross-presentation of microbial antigens to CD8+ lymphocytes via MHC Class I pathways. The goals of the current proposal are first, to characterize the cellular and cytokine responses in the lung observed early after infection with SOD-diminished M. tuberculosis, as rapid pulmonary interstitial infiltration with mononuclear cells undergoing apoptosis appears to be a process unique to the SOD-diminished strains that is not observed during infection with either virulent M. tuberculosis or the current vaccine strain for tuberculosis, BCG. This should define the conditions under which antigen cross-presentation occurs in vivo, yielding information that may be useful for a variety of vaccines. The second goal is to construct non-reverting SOD-diminished mutants of H37Rv and BCG by replacing the wild-type SOD allele with mutant alleles, some of which encode enzymatically less efficient mutants of SOD. This should yield a SOD-diminished vaccine candidate that is stable and safe enough for administration to man. The third goal is to determine the optimal level of SOD production for maximum vaccine efficacy and the immune correlates of protection. Diminishing the production of factors produced by intracellular pathogens that inhibit macrophage apoptosis is a strategy for making new vaccines that achieve MHC Class I antigen presentation. This should have implications not only for tuberculosis but for other infectious diseases in which CD8+ T-cell responses are a critical component of a protective immune response.